292
J. Med. Chem. 1984,27, 292-301
complex side chains as well as to create further structural variation at the level of two- and three-carbon side-chain substituents.
zoek (Grant 30048.75), and by the Belgian Geconcerteerde Onderzoeksacties (Grant 81/86-27). We thank Anita Van Lierde for her dedicated technical assistance.
Acknowledgment. This work was supported by the
Registry No. 1, 69-33-0; 2, 24386-95-6; 3, 21193-80-6; 4, 24386-93-4; 5,78000-56-3;6,78000-57-4; 7,78000-55-2;8,606-58-6; 9,18417-89-5; io, 76319-88-6; 11,76319-87-4; 12,76319-84-1; 13, 76319-82-9; 14, 87984-10-9; 15, 87938-37-2; 16, 87938-38-3; 17, 58316-88-4; 18, 36791-04-5; 19, 54262-83-8.
U.S.Army Medical Research and Development Command under Research Contract DAMD-17-79-C-8014, by the National Cancer Institute (Grant CA 21493), by the Belgian Fonds voor Geneeskundig Wetenschappelijk Onder-
Pyridonecarboxylic Acids as Antibacterial Agents. 2.’ Synthesis and Structure-Activity Relationships of 1,6,7-Trisubstituted 1,4-Dihydro-4-oxo-1,8-naphthyridine-3-carboxylic Acids, Including Enoxacin, a New Antibacterial Agent2 Jun-ichi Matsumoto,* Teruyuki Miyamoto, Akira Minamida, Yoshiro Nishimura, Hiroshi Egawa, and Haruki Nishimura Research Laboratories, Dainippon Pharmaceutical Co., Ltd., Enoki 33-94, Suita, Osaka, 564, Japan. Received May 19, 1983 The title compounds having nitro, amino, cyano, chloro, or fluoro as the C-6 substituent were prepared. Introduction of the chloro and cyano groups at C-6 was accomplished by the Sandmeyer reaction of 6-amino-1,8-naphthyridine derivatives 9 via their 6-diazonium salts 10. The reaction was extended to the synthesis of the 6-fluor0 analogues 20, involving the Balz-Schiemann reaction of the diazonium tetrafluoroborate 19. Furthermore, a series of the 1-ethyl (24 and 27-30), 1-vinyl (35-37), l-(2-fluoroethyl) (38 and 39), and 1-(difluoromethyl) (40) analogues of 7-substituted 6-fluoro-l,4-dihydro-4-oxo-l,8-naphthyridine-3-carboxylic acids was prepared. 1-Pyrrolidinyl and, particularly, N-substituted or unsubstituted 1-piperazinyl groups were introduced as the C-7 variants. As a result of this study, l-ethyl-6-fluoro-1,4-dihydrc~4-oxo-7-( l-piperazinyl)-l,&naphthyridine-3-~boxylic acid (24b named enoxacin, originally AT-2266) was found to show the most broad and potent in vitro antibacterial activity, an excellent in vivo efficacy on systemic infections, and a weak acute toxicity. Structure-activity relationships of compounds with variations of substituents at C-1, C-6, and C-7 are also discussed.
In earlier papers, we had reported the synthesis and antibacterial activity of pyrido[2,3-d]pyrimidines(1):” 1,6and l,&naphthyridines (2 and 3),l and quinolines (4),5 all of which possess a 1,4-dihydro-4-oxopyridine-3-carboxylic acid moiety as a common structure. This class of compounds has increasingly attracted attention as a source of new antibacterial agentsq6 It has been reported that nalidixic acid (3, R1 = CH,; R2 = C2H6)7and oxolinic acid [4, R1 = 6,7-(-OCH20-),R2 = C&]’ are a specific inhibitor
Chart Ia **cOzH
R
RZ
a
(1) Paper 1: Hirose, T.; Mishio, S.; Matsumoto, J.; Minami, S. Chem. Pharm. Bull. 1982.30. 2399.
(2) This work was presented in p k (a) at the 11th International Congress of Chemotherapy and 19th Interscience Conference on Antimicrobial Agents and Chemotherapy, Boston, MA, Oct 1979 (In “Current Chemotherapy and Infectious Disease”; Nelson, J. D.; Grassi, C., Eds.; American Society for Microbiology: Washington, DC, 1980; Vol. 1, p 454) and (b) at the 100th Annual Meeting of the Pharmaceutical Society of Japan, Tokyo, Apr 1980; Abstr, p 275. (c) Matsumoto, J.; Takase, Y.; Nishimura, Y. European Patent 9425,1980; Chem. Abstr. 1980, 93, 168305~. (3) Minami, S.; Shono, T.; Matsumoto. J. Chem. Pharm. Bull. 1971,19, 1426. (4) Matsumoto. J.: Minami. S. J.Med. Chem. 1975. 18. 74. (5) Minami, S.iMatsumoto; J.; Sugita, M.; Shimizu,’M.; Takase, Y. German Offen. 2362553, 1974; Chem. Abstr. 1974, 81, 105562k. (6) For a review, see Albrecht, R. Prog. Drug Res. 1977, 21, 9. (7) Lesher, G. Y.; Froelich, E. J.; Gruett, M. D.; Bailey, J. H.; Brundage, R. P. J. Med. Pharm. Chem. 1962,5, 1063. (8) Kaminsky, D.; Meltzer, R. I. J. Med. Chem. 1968, 11, 160. 0022-2623184J 1827-0292$01.50/0
.ABI
l,A=B=N 4, A = B = CH 5, A = CR3;B = N 2, A = N ; B = CH 6, A = C F ; B = C H ; R Z = C,H, 3,A = C H ; B = N a, R’ = b, R’ = H N ~ N - ; c, R’ = cwJ--u A W Wv-
0-;
of DNA synthesis in susceptible bacterial cells, and this inhibition is due to the prevention of DNA gyrase activity? Recent findings indicate that piromidic acid (la, R2 = C2H5)3and pipemidic acid (lb, R2 = C2H6)4also act by the same mechanism.1° A continuing interest in this field led us to an investigation of 1,6,7-trisubstituted 1,4-dihydro-4-oxo-l,8naphthyridine-3-carboxylicacids (5), with the C-6 substituent being either nitro, amino, chloro, or fluoro; little information about the effect of the C-6 substituent upon antibacterial activity has been available thus far. ~
~~
(9) For a review, see: Crumplin, G. C.; Midgley, J. M.; Smith, J. T. “Topics in Antibiotic Chemistry”; Sammes, P. G., Ed.; Ellis Horwood: West Sussex, England, 1980; Vol. 3, pp 11-38. (10) Yamagishi, J.; Furutani, Y.; Inoue, S.; Ohue, T.; Nakamura, S.; Shimizu, M. J.Bacteriol. 1981, 148, 450. 0 1984 American Chemical Society
1,4-Dihydro-4-oxo-l,8-naphthyridine-3-carboxylic Acids
Scheme II@
Scheme I a R2&
C02C2H5
R'
Journal of Medicinal Chemistry,1984, Vol. 27, No. 3 293
-
I
I
'ZH5
C2H 5
13
7,R2 = H 8, R 2 = NO, 9 , R 2 =NH,
R
R
I
CZH 5
c2H5 I
C2H5
l l a , R 2 = C1; R3 = C2H, b, RZ = CN; R3 = C,H, c , R 2 = F ; R 3 =C,H, 1 2 a , R * = C1; R3 = H b, R Z = CN; R3 = H
l o a , X = C1 b, X = CN C , X = BF,
a
For 7-9:
a, R' = CH,O; b, R'
c, R' = G - ( e x c e p t
=CH~C,"~N-
W
llb
-
14
15
POCIS
R
;
for 7 ) .
I
C2H5
C2H5
16 17, R 2 = C,H, 1-Pyrrolidinyl, 1-piperazinyl, and N-methyl-l18. R 2 = H piperazinyl groups were selected to be introduced at C-7 a For 15, 17, and 1 8 : a, R' =o-; R1 b =~~r'hl-; , of 5 at the outset of this study on the basis of our successful n W development of piromidic and pipemidic acids. As a result c, R' =CH3--d k-. of the present study, we found a promising candidate, W l-ethyl-6-fluoro-1,4-dihydro-4-oxo-7(l-piperazinyl)-l,& in good yield by diazotization of 9a with sodium nitrite in naphthyridine-3-carboxylicacid (24b), as a potent new a dilute sulfuric acid, followed by successive treatment with antibacterial agent. Recent papers dealing with the a mixture of cuprous cyanide and potassium cyanide. The quinoline analogues norfloxacin (6b)ll and perfloxacin esters lla,b were hydrolyzed to the acids 12a,b, respec(6c)12prompted us to report our results on a synthesis of tively. the 1,B-naphthyridinederivatives 5, as well as 24b, and the Compounds lla,b served as the starting compounds for structure-activity relationships (SARs) associated with the preparation of the 6-chloro and 6-cyano derivatives 15 variations of the C-1, (2-6, and C-7 substituents of 5. and 18, respectively, with a cyclic amino group at C-7 Although a number of l,&naphthyridine derivatives 3 (Scheme 11). Alkaline hydrolysis of lla, followed by without the C-6 substituent have been reported, little chlorination of 13 with phosphoryl chloride, afforded the appears to be known about derivatives of 5 with substit6,7-dichloro compound 14. Regiospecific displacement of uents at C-616,13probably due to synthetic difficulties. An the 7-chloro group in 14 with the selected amine was efattractive intermediate for the synthesis of 5 would be a fected by refluxing in acetonitrile to give good to excellent 6-diazonium salt of an appropriately functionallized yields of the desired compounds 15a-c; in this case, the naphthyridine, such as 10, which might be converted into possibility of an isomeric structure due to displacement the 6-cyano, 6-chloro, and 6-fluor0 analogues 11 under of the 6-chloro group by the amine was excluded because conditions of the Sandmeyer reaction or the like as dethe product 15a was identical with a compound that was picted in Scheme I. derived through an unambiguous route involving the reChemistry. The requisite diazonium salt 10 was action of 12a with pyrrolidine. Treatment of the 6readily prepared by diazotization of the corresponding cyano-7-methoxy compound 1l b with phosphoryl chloride 6-amino compound 9 via the 6-nitro analogue 8. Thus, ethyl l-ethyl-1,4-dihydro-7-methoxy-4-oxo-1,8- gave directly the 6-cyano-7-chloroderivative 16, which was then treated with the amines in acetonitrile to afford the naphthyridine-3-carboxylate(7a) was nitrated with a 6-cyano-7-(cyclic amino) esters 17a-c. These esters were mixed acid (fuming nitric acid and concentrated sulfuric converted into the corresponding carboxylic acids 18a-c acid) to give 8a in 81% yield. Reduction of 8a with iron by acidic hydrolysis. powder in acetic acid gave the 6-amino derivative 9a, which In order to introduce a fluoro group at C-6 of the 1,8was then diazotized with sodium nitrite in concentrated naphthyridine ring, the Balz-Schiemann reaction14 was hydrochloric acid. The resulting diazonium chloride loa, applied initially to the 6-amino-7-methoxy derivative 9a. without isolation, was successively treated with cuprous Diazotization of 9a with sodium nitrite in 42% tetrachloride, giving the 6-chloro compound lla in 64% yield. fluoroboric acid gave an 85% yield of the diazonium tetThe 6-cyano congener l l b could analogously be prepared rafluoroborate lOc, which showed a characteristic IR absorption at 2200 cm-l (v, NEN). However, attempts to (11) Koga, H.; Itoh, A.; Murayama, S.;Suzue, S.; Irikura, T. J. Med. convert the diazonium salt 1Oc into the fluoro compound Chem. 1980,23, 1358. 1ICunder a variety of conditions were unsuccessful. (12) Goueffon, Y.;Montay, G.;Roquet, F.; Pesson, M. C. R. Hebd. ~
~~
~
Seances Acad. Sci. 1981,37. (13) (a) Suzuki, N.; Kato, M.; Dohmori, R. Yakugaku Zasshi 1979, 99, 155. (b) Suzuki, N. Chem. Pharm. Bull. 1980,28, 761.
(14) Roe, A. Org. React. 1949,5,193-228.
294 Journal of Medicinal Chemistry, 1984, Vol. 27, No. 3
Matsumoto et al.
Table I. l-Ethyl-l,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic Acid Derivatives yield,a compd
mp, "C
8a 8b 8c 9a 9b 9c 1l a llb 12a 12b 13 14 15a
recrystn solvent
224-221 23 1-212 205-201 250-252 265-210 210-21 2 e 156-157 221-223 211-215 281-284 >300 26 2-261 29l-30Og
%
synth method
formulaC
EtOH EtOH EtOH EtOH EtOH EtOH EtOH-H,O EtOH DMF DMF DMF DMF DMF
80.8 A C14H15N306 89.5 A C19H23N506 91.6 B C17H20N405 19.9 C C14H17N304 78.2 C C19H25NS04 31.5 C C17HZ2N403 D 63.7 C14H15C1N204 E 66.0 C15H1SN304 F 12.3 12 H l 1 'IN 2 4 F 93.0 C13H11N304 G 83.3 '11 H9C1N204 H 65.7 11H8C12N20 3 I 85.1 15 H16 3' J 75.6 15bh 246-248 AcOH/NH40H I 82.1 1S H 1 4 3 15C I 232-233 DMF 11.6 C16H19C1N403 2 5 2-2 56 16 77.1 DMF H C 1 4 H l ZCIN ' 3 3 17a 259-260 75.3 EtOH K C18H20N403 151-1 54 17b 51.7 DMF K C,,H,,N5O3~1.2H,O 1 95-1 96 17c EtOH 14.2 K C19H23N503 296-298 18a 91.1 DMF F C16H16N403 18b F 242-245 dec HCl/AcONa' 7914 C16H17N503 233-235 18c DMF F 13.6 C17H19N503 20a EtOH-CH,CI, 22 0-2 22 8.2 L C17H20FN 3' 84.7 M 20b 19 5-1 91 AcOEt 36.2 L 89. I M 22a 256-258 95.1 F DMF C15H16N405 22b 234-236 DMF N 80.9 C15H17N505'2H20 22c 219-221 E tO H-CHCl 54.9 R C16H,,N,05~0.3H,0 23a 290-296 dec NaOH/AcOH' 85.1 0 C15H18N403 253-255 23b NaOH/AcOH N 29.3 C15H19N503'2H20 23c EtOH 2 53- 2 56 P 26.1 C16H,1 N5° 3 24a AcOEt-CH2C12 29 9-300 F 13.5 ClSH16FN 3' 24b' EtOH-CH,Cl, 15.3 220-224 Q C15H17FN403 EtOH-CH,Cl, 24c R 15.0 221-230 C16H19FN403 T_____-___ ______ 89.3 -. a Yields are of purified products and are not maximal. Capital letters refer to the method described in the Experimental Section. All compounds were analyzed for C, H, N,and, where present, C1 and F ; analytical results were within +0.3% of the theoretical values. Literature13amp 200-201 "C. e Literature13a mp 259 "C. L i t e r a t ~ r e mp f ~ ~270-213 "C. Literature13amp >300 "C. HCI salt: mp 290-300 "C. Anal. (C,,H,,C1N40;HC1) C, H, C1, N. Purified by reprecipitation on treatment with the acid and subsequently with the base or vice versa. Literature13bm1) 243 "C. HCl salt: mp 265-273 "C. Anal. (Cl6H,,N~O;HC1)C , H, C1, N . l For salts, see the Experimental Section. J
J
Scheme 111'
Chart 11'
( I2) ) NaN02, HPFS HCI
-
U
I/
c
9b9c
,.
pF6-N2m 0
C02C2H5
R
I I
C2H5
19
22, R2 = NO, 23,RZ =NH, 24, R Z = F
1hed
a
A - cH3-NuN .
a,R1 = o - ; b , R 1 =wA~--;c,R1 W
Diazotization of 9b with sodium nitrite in 65% hexafluorophosphoric acid furnished a 94% yield of the 6-diazonium hexafluorophosphate 19b (Scheme 111). When 21 19b was heated at 80-100 "C with n-heptane, the fluorination proceeded smoothly to give a 36% yield of the 20 desired 6-fluor0 compound 20b, though unsatisfactory in a For 19-21: a, R =(-)-; b, R = yield. The same conditions permitted the conversion of 7-pyrrolidinyl-6-diazonium hexafluorophosphate 19a into the 6-fluoro-7-pyrrolidinylanalogue 20a but in a very poor Compound 9b was then employed for the Balz-Schiemyield. Compounds 20a,b were alternatively prepared in ann reaction. Nitration of ethyl 7-(4-acetyl-lpiperaziny1)-1-ethyl-1,4-dihydr0-4-oxo-l,S-naphthyridine- good yields by the N-ethylation of 21a,b, respectively. The synthesis of 21, which is important for aspects of the 3-carboxylate (7b), followed by reduction of the 6-nitro large-scale preparation of 20 and its derivatives, particucompound 8b, gave the requisite amino analogue 9b.
Journal of Medicinal Chemistry, 1984, Vol. 27, No. 3 295
1,4-Dihydro-4-oxo-1,8-naphthyridine-3-carboxylic Acids
Table 11. l-Ethyl-6-fluoro-1,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic Acids Derivatives U
I
'ZH5
compd
R
mp, "C
yield:
recrystn solvent
%
>300 DMF 27a 2" 273-276 ACOHINH~OH~ 27b NH,CH,CH,NH 27c 3-oxo-1-piperazinyl >300 DMF 27d CH,(CH,CH,),N211-213 DMF 261-263 DMF 27e O(CH,CH,),N251-253 MeCN 27f S( CH,CH,),NA~OHINH~OH~ 238-240 27g homopiperazinyl e EtOH 192-193 27h 1-azepinyl EtOH 27i CH,( CH,CH,CH,),N188-190 MeCN 237-239 27j Ph-N( CH,CH,),NEtOH-CH,Cl, 220-222 27k PhCH,N( CH,CH,),NEtOH-CH,Cl, 193-194 271 Et-N(CH,CH,),NEtOH-CH,Cl, 208-209 2 8a ~ - P N( P CH ,CH ,) ,NEtOH-CH,Cl, 28b n-Bu-N( CH,CH,),N184-185 213-214 E tOH-CH,Cl, 2 8 ~ s-Bu-N(CH,CH,),N292-294 EtOH-CH,Cl, 29 OHC-N( CH,CH,),N30 CH,CON(CH,CH,),N> 300 E t OH-C H ,C1, __Purified by reprecipitation on treatment with See footnotes a-c in Table I. base. e Hexahydro-lH-1,4-diazepinyl.
synth methodb
formulaC
83.3 S C11H10FN303 83.4 T C13H15FN403 71.4 T M H 1SF "3'4 84.3 T C16H18FN303 79.6 T Ci Pi6FN-04 83.5 T 33'' 72.7 T C16H1YFN403 74.0 T C,,H,'lFN303 78.1 T C18HZZFN 3' 50.4 T C21H21FN403 31.7 T C22H23FN403 91.6 T C1,H21FN403 32.4 U C18H23FN403 61.2 U C19H25FN403 26.5 U lYH XFNdO 3 97.5 V 16H 17FN404 85.6 0 C17H1YFN404 the acid and subsequently with the
Table 111. 1,7-Disubstituted 6-Fluoro-1,4-dihydro-4-oxo-1,8-naphthyridine-3-carboxylic Acid Derivatives yield,a synth compd mp, "C recrystn solvent % method formula M 213-215 MeCN 31 92.9 19 H23FN40 5 90.3 32 168-170 AcOEt C1YH22C1FN404 M 184-185 Me,CO 79.8 33 ClYH*,F,N4O4 AcOEt-CH,Cl, X 282-284 19.4 34 C18H1YF3N404 MeCN Y 264-266 35 80.6 C17H17FN404 AcOH/NH40He 256-260 dec Z 36 82.4 C15H15FN403 205-207 MeCN R 43.6 37 C16H1,FN403 223-225 HCl/NH40He 38 75.4 Q Cl,H,, FZN.40 3 224-226 MeCN 39 R 82.2 C16H18F2N403 DMF-EtOH 40 284-286 97.6 Q C,4H,xF,N40% See footnotes a-c in Table I. HC1 salt: mp 290 "C dec (EtOH-H,O). Anal. (Cl,H,5FN,03~HCl) C, H, C1, F, N. Methanesulfonic acid salt: mp 292-294 "C dec (EtOH-H,O). Anal. (Cl,HlsFN,03~CH4S031/,H,0) C, H, F, N, S. Purified by reprecipitation on treatment with the acid and subsequently with the base.
w
-
larly 24b, will be reported e1~ewhere.I~ The 6-fluor0 esters 20a,b thus obtained were hydrolyzed under acidic conditions into the corresponding 6-fluor0 carboxylic acids 24a,b. Reductive N-methylation of 24b upon treating with formaldehyde and formic acid afforded the N-methylpiperazinyl compound 24c. Analogously, the 6-nitro carboxylic acids 22a,b were prepared by hydrolysis of 8c,d, respectively, and 22c was derived from 22b by the reductive N-methylation. The 6-amino carboxylic acids 23a-c were obtained by reduction of the corresponding nitro compounds 22a-c. Since compound 24b showed an excellent in vitro antibacterial activity as described in the following section, the 6-fluor0 analogues substituted at C-7 by cyclic (and acyclic) amino groups other than the piperazinyl group of 24b were prepared (Scheme IV). Prolonged heating of 20b with alkali, followed by chlorination of 25 with phosphoryl chloride, afforded the 7-chloro-6-fluoroderivative 26. Upon treatment of 26 with a variety of amines, the displacement reaction proceeded quite regioselectively (15) Matsumoto, J.; Miyamoto, T.; Minamida, A,; Nishimura, y.; Egawa, H.; Nishimura, H.accepted for publication in J. Het-
erocycl. Chem. See also ref 2c.
Scheme IV 24b
20b
for 20 and 29
Fmco2H Imco ,.
h
U
U
I1
R'
II
*f
I
I
'ZH5
CZH 5
25, R' = HO 26, R' = C1
27-30 (Table V )
at C-7 to give the desired 7-substituted 6-fluor0 derivatives 27a-1. Reductive alkylation on the piperazinyl N-4 of 24b was carried out by treatment with propionaldehyde, butyraldehyde, and isobutyraldehyde in formic acid to afford the corresponding N4-alkyl derivatives 28a-c. N-Formyl compound 29 was prepared by treatment of 24b with formamide in formic acid. Partial hydrolysis of 20b by alkali gave the N4-aCetYlcarboxylic acid 30. Among these compounds thus prepared, 28a-c, 29, and 30 were identified as metabolites of 24b in animals tested.16
296 Journal of Medicinal Chemistry, 1984, Vol. 27, No. 3
Scheme V
Matsumoto et al.
Table IV. In Vitro Antibacterial Activitya _.__---
21b
-
F.n& C02C2H5
CH,CONn
u
I R
31, R = CH,CH,OH 32, R = CH,CH,Cl 33, R = CH,CH,F 34, R = CHF,
35, R = CH=CH,; R' = CH,CO 36, R = CH=CH,; R' = H 37, R = CH=CH,; R' = CH, 38, R = CH,CH,F; R' = H 39, R = CH,CH,F; R' = CH, 40, R = CHF,; R' = H
Finally, the ethyl group at N-1 of 24b and 24c was replaced by a vinyl, fluoroethyl, or difluoromethyl group (Scheme V), because these groups seemed attractive as variants that might possibly improve the antibacterial activity of the pyridonecarboxylicacids.l' Thus, treatment of 21b with ethylenechlorohydrin,followed by chlorination of 31 with thionyl chloride, afforded the chloroethyl derivative 32. On treatment of 32 with potassium hydroxide in ethanol, elimination of the hydrogen chloride proceeded smoothly to give an 81% yield of the vinyl compound 35, along with a small amount of the hydrolyzed product, 6-fluoro-l,4-dihydro-7-hydroxy-4-oxo-l-vinyl-l,8naphthyridine-3-carboxylicacid (41). The N4-protective group in 35 was removed by alkaline hydrolysis to give the desired piperazinyl vinyl derivative 36, which was subsequently heated with 37% formalin in formic acid to produce the N4-methylpiperazinyl vinyl analogue 37. Furthermore, the fluoroethyl derivative 33 was prepared from 21b by the reaction with 2-fluoroethylp-toluenesulfonate. Subsequent acid treatment of 33 provided the deprotected acid 38, which was converted into the N4-methylpiperazinyl analogue 39 by reductive methylation. The difluoromethyl compound 40 was prepared, though in a poor yield, by the reaction of 21b with (difluoromethyl)carbene, formed in situ from sodium chlorodifluoroacetate, followed by deprotection of 34 with hydrochloric acid. Biological Results and Discussion The in vitro antibacterial activity of compounds prepared in the present study against representatives of Gram-positive (Staphylococcus aureus 209P JC-1) and Gram-negative bacteria (Escherichia coli NIHJ JC-2 and Pseudomonas aeruginosa Tsuchijima) is shown in Table IV. Minimal inhibitory concentrations (MIC's) of 3a-c (R2 = CZH5),lnalidixic acid, and pipemidic acid are included for comparison. (16) Nakamura, R.; Yamaguchi, T.; Sekine, Y.; Hashimoto, M. J. Chromatogr. 1983, 278, 321. (17) For example, see ref 4. Albrecht, R. Eur. J.Med. Chem. 1977, 12, 231. Agui, H.; Saji, I.; Nakagome, T. Japanese Patent Kokai 139094,1977; Chem. Abstr. l978,88,105406g. Lee, K. T. German Offen. 2706781, 1977; Chem. Abstr. 1977, 87, 201507r.
compd
min inhibitory concn, wg/mL S. aureus E. coli P. aeruginosa 209P JC-1 NIHJ JC-2 Tsuchijima
>loo 3a 12.5 25 6.25 25 25 3b 25 6.25 25 3c >loo > 100 12.5 15a 0.78 6.25 3.13 15b 1.56 12.5 15c 6.25 25 12.5 3.13 18a 6.25 1.56 6.25 18b 12.5 1.56 12.5 18c >loo 100 25 22a 25 6.25 6.25 22b 50 3.13 12.5 2 2c >loo > 100 > 100 23a 6.25 > 100 3.13 23b 12.5 25 1.56 23c 3.13 1.56 0.39 24a 0.78 0.2 0.78 24b 1.56 0.39 1.56 24c 1.56 50 >loo 27a 50 6.25 25 27b 12.5 6.25 0.78 27c 12.5 0.78 6.25 27d 6.25 3.13 27e 1.56 12.5 1.56 3.13 27f 0.78 1.56 1.56 27g 6.25 50 3.13 27h > 100 12.5 25 27i 12.5 >loo 6.25 27j 6.25 12.5 0.78 27k 3.13 0.78 2 71 0.78 1.56 12.5 1.56 2 8a 25 3.13 3.13 28b 3.13 25 1.56 2 8c 1.56 12.5 3.13 29 50 3.13 3.13 30 0.1 0.2 1.56 36 0.78 0.2 3.13 37 0.78 0.2 0.39 38 1.56 0.2 0.78 39 0.78 6.25 6.25 40 50 N A ~ 50 1.56 6.25 1.56 PPA 6.25 Nalidixic acid. a See the Experimental Section. Pipemidic acid. Firstly, we wish to discuss the SAR's associated with modification of the C-6 substituent of compounds 15, 18, and 22-24 compared with 3a-c (R2 = C,H,). Introduction of an amino, nitro, cyano, chloro, or fluoro group at C-6 of 3a-c influenced markedly the antibacterial activity. Thus, in a series of the pyrrolidinyl compounds (15a, 18a, 22a, 23a, and 24a), the fluoro and cyano groups cause an increase in activity against all the bacteria tested, whereas others result in a complete loss of activity, particularly against the Gram-negative bacteria; it is noticed that a remarkable difference between the fluoro and the chloro groups in an effect upon the activity is observed in the series a. With respect to the piperazinyl and N-methylpiperazinyl derivatives (i.e., series b and c of compounds 15, 18, and 22-24), introduction of the C-6 substituent tends to enhance the activity against both Gram-positive and Gram-negative organisms, with a few exceptions. In both series of compounds, the activity against S. aureus increases in the order NH, 5 H < NO2 = CN < C1 < F, whereas the Gram-negative activity follows the sequence NOz IH < NH2 = CN = C1 < F, with the amino, cyano, and chloro compounds being equally active. In the b series of compounds, the activity against Escherichia coli increases in the order H = NOz < NH2 < CN < C1 < F. It is of particular interest that replacement of hydrogen by
1,4-Dihydro-4-oxo-l,8-naphthyridine-3-carboxylic Acids
Journal
of Medicinal Chemistry, 1984,
Vol. 27, No. 3 297
Table V . Efficacy on Systemic Infections and Acute Toxicity with Oral Administration in Micea
e
compd 24b 24c 36 37 38 39 N A ~ PPA e
S. aureus 50774: ED,,b (MIC)' 1 0 (0.78) 4.8 (1.56) 33.4 (3.13) 10.5 (3.13) 11.5 (1.56) 1.4 (0.39) > 800 ( 50) 215 (25)
a See the Experimental Section. Pipemidic acid.
E. coli P-5101: EDSO WIG) 1.8 (0.1) 1.2 (3.13) 1.3 (0.1) 1.1(0.1) 3.0 (0.1) 0.52 (0.1) 29.2 (3.13) 12.8 (3.13)
In millimams Der kilogram.
halogen, especially fluorine, at C-6 in the 1,haphthyridine system results in an outstanding enhancement of the activity against both Gram-positive and Gram-negative organisms. A comparison of the activity of the piperazinyl (15b, 18b, 22b, 23b, and 24b) and the N-methyl-lpiperazinyl analogues (15c, 18c, 22c, 23c, and 24c) with the same C-6 substituent generally found the former to be more active than the latter, except for the activity of two sets (22b vs. 22c and 24b vs. 24c) against E . coli. Compound 24a, bearing the pyrrolidine ring at C-7, is the most potent as far as activity against S. aureus, whereas 24b and 24c, bearing the piperazine ring instead, have better activity against Gram-negative organisms. Overall, the most active is compound 24b, which is about 30 times as active as 3b (R2= C,H5) against every organism tested. SAR's of the C-7 substituent in a series of analogues with an ethyl group at N-1 (see 24 and 27-30) were substantially comparable to those for the corresponding 5,8-dihydro-5oxopyrido[2,3-d]pyrimidine-6-carboxylicacids (1, R2 = C2H5),which had previously been studied by ~ 9 . ~Thus, 9 ~ modification of the cyclic amino moiety at C-7 resulted in a significant decrease in activity as observed in 24a and 27c-i compared with 24b. Only compound 24a with the 1-pyrrolidinyl group, however, is more active than 24b against the Gram-positive Staphylocococcus strain. A comparison of the activity between 24b and 27d, as well as 27g and 27h, obviously indicates that the presence of a basic NH group in the cyclic amino function is a prerequisite to optimal activity. However, if this NH group is an amide, such as in the 3-oxo-1-piperazinylgroup of 27c, activity is decreased. Introduction of an alkyl, aralkyl, or aryl group a t the piperazinyl N-4 of 24b (Le., 24c, 27j-1, and 28a-c) causes a decrease in activity; in particular, the activity against Gram-negative bacteria is reduced markedly with an increase in the chain length and, hence, with an increase in the lipophilicity of the group introduced. Compounds 27a-c, 29, and 30, identified as metabolites of 24b,16are substantially less active. Of these derivatives possessing the ethyl group at N-1, no compounds superior to 24b in activity were found. Variation of the N-1 substituent on the naphthyridine ring significantly influenced the in vitro activity. The excellent activity of the vinyl and fluoroethyl analogues (see 36-39) should be noted. The difluoromethyl group in 40, however, led to a considerable decrease in activity. In each comparison between 24b and 36, as well as 24c and 37, it is revealed that replacement of the ethyl group by a vinyl group causes an increase in the Gram-negative activity, whereas it reduces appreciably the Gram-positive activity. The fluoroethyl function, in contrast to the vinyl group, enhances the Gram-positive activity, while the Gram-negative activity is essentially unchanged (compare 24b and 24c with 38 and 39, respectively). Selected compounds (24b,c and 36-39) were then tested on systemic infections due to S. aureus 50774, E. coli
P. aeruginosa 12: ED,, (MIC) 9.0 (0.78) 10.6 (0.2) 2.4 (0.39) 3.7 (1.56) 27.2 (0.78) 4.2 (1.56) 267 (100) 70.8 (25)
In micrograms per milliliter.
LD 50 9 mglkg PO >2000 210 >2000 354 > 2000 1866 1800 >2000 Nalidixic acid.
P-5101, and P. aeruginosa 12, with oral administration in mice, and compared with nalidixic acid and pipemidic acid. The results are listed in Table V, which includes for reference the MIC's against the organisms employed. The median effective dose (ED,,) values of these compounds are always much lower than those of the reference drugs. With the exception of 38, the in vivo efficacy on the experimental infection due to the Gram-negative bacteria was highly increased by changing the N-1 substituent from ethyl to vinyl or fluoroethyl (see 36, 37, and 39). When the ethyl group of 24b was changed to fluoroethyl (38), however, the efficacy on the pseudomonal infection was reduced to one-third the potency of the parent compound. On the other hand, this variation at N-1 resulted in a decreased efficacy on the infection due to S. aureus, a Gram-positive bacterium; for example, the most effective compound, 36, on the pseudomonal infection is the least effective on the staphylococcal infection. Of much interest is compound 39, which is about 7, 3.5, and 2 times as potent as 24b on systemic infections caused by S. aureus, E. coli, and P. aeruginosa, respectively. However, an acute toxicity test with a single oral administration in mice revealed the most effective compound, 39, to possess a smaller value of the median lethal dose (LD5,,) than that of 24b, but it is comparable to the LD50 of nalidixic acid, as shown in Table V. It is noteworthy that compounds 24c and 37, as well as 39, which have the N4-methyl group on the piperazine ring in common, exhibit more potent toxicity than the corresponding N4-unsubstituted compounds 24b, 36, and 38, of which every LD50 is >2000 mg/kg orally in mice. As a result, compound 24b (named enoxacin, originally AT-2266) was found to possess broad and potent in vitro and in vivo antibacterial activity and weak oral acute toxicity. These findings indicate the possible use of 24b as a potent, orally administrable antibacterial agent. Enoxacin (24b) was thus selected for further biological evaluation1*and is now undergoing clinical trials. Experimental Section Chemistry. All melting points were determined on a Yanagimoto micromelting point apparatus and are uncorrected. IR spectra were recorded on a Hitachi Model 215 spectrophotometer. 'H NMR spectra were taken at 60 or 100 MHz on either a Varian EM-360A or HA-100D spectrometer with Me4Si as an internal standard. Mass spectra were recorded on a Hitachi RMU-6L single-focusing mass spectrometer using the direct inlet system at 70-eV ionization potential. Elemental analysis are indicated only by symbols of the elements; analytical results were within *0.3% of theoretical values. IR, lH NMR, and/or mass spectra (18) Nakamura, S.; Minami, A.; Katae, H.; Inoue, S.; Yamagishi, J.; Takase, Y.; Shimizu, M. Antirnicrob. Agents Chernother. 1983, 23, 641. Nakamura, S.; Nakata, K.; Katae, H.; Minami, A.;
Kashimoto, S.; Yamagishi, J.; Takase, Y.; Shimizu, M. Ibid. 1983, 23, 742.
298 Journal of Medicinal Chemistry, 1984, Vol. 27, No. 3
Matsumoto et al.
were obtained on all compounds and were consistent with assigned ane-acetone to give 0.58 g (41.0%) of 7a;this was identical in all structures. Organic solutions were dried over anhydrous Na2S04 respects with an authentic specimen of 7a. or MgS04. Method D. Ethyl 6-Chloro-l-ethyl-1,4-dihydro-7-methEthyl 1-Ethyl-l,l-dihydr0-7-met hoxy-4-oxo-1 3 oxy-4-oxo-l,8-naphthyridine-3-carboxylate (lla). To a stirred naphthyridine-3-carboxylate (7a). In a similar procedure to solution of Sa (4.0g, 13.7 mmol) in 50 mL of concentrated HCl that described in our previous paper,' 7a was prepared from was added dropwise a solution of NaNOP (1.5g, 21.7 mmol) in 2-amino-6-methoxypyridineby the Gould-Jacobs-type reaction 8 mL of water over a period of 10 min at 0-3 "C. After an of its malonate, followed by the N-ethylation of ethyl 1,4-diadditional 10 min, the reaction mixture was added to a solution hydro-7-methoxy-4-oxo-1,8-naphthyridine-3-carboxylate; the of cuprous chloride (2.7g, 13.6mmol) in 15 mL of concentrated details will be reported elsewhere:le mp 142.5-143 "C; IR (KBr) HC1 and allowed to stir at room temperature for 1 h, after which 1680,1640cm-l; EIMS, m / z 276 (M'), 240 (M' - C2H4CO2).Anal. time the temperature was maintained at 65 "C for 30 min. The mixture was neutralized with 10% NaOH and extracted with ( C I ~ H I ~ N PCO y ~H,) N. Ethyl 7-(4-Acetyl-l-piperazinyl)-l-ethyl-1,4-dihydro-4- CHC1,. The extract was concentrated to dryness in vacuo; the oxo-l,8-naphthyridine-3-carboxylate (7b). A mixture conresidue was crystallized from aqueous EtOH to give 2.71 g (63.7%) taining 5.0 g (17.8mmol) of ethyl 7-chloro-l-ethyl-1,4-dihydro- of lla: IR (KBr) 1720,1690,1620em-'; EIMS, m / z 310 (M+), 4-oxo-1,8-naphthyridine-3-carboxylate,1 7.0 g (54.7 mmol) of 238 (M' - C2HdCO2); 'H NMR (CDC13) 6 8.70 (1 H, 9, H-5), 8.60 N-acetylpiperazine, and 100 mL of EtOH was heated to reflux (1 H, 8, H-2), 4.45 (4H, q, J = 7 Hz, NCHZCH,, COZCHZCH,), for 3 h. The reaction mixture was concentrated to dryness in 4.20 (3 H, 8, OCHB), 1.55 (3 H, t, J = 7 Hz, CO,CH,CH,), 1.44 vacuo. The residue was taken up in 80 mL of water, acidified (3 H, t, J = 7 Hz, NCHZCH,). with 10% HCl, and extracted with CHC13. The extract was Method E. Ethyl 6-Cyano-l-ethyl-1,4-dihydro-7-methwashed with water and dried, and the solvent was evaporated in oxy-4-oxo-1,8-naphthyridine-3-carboxylate (1 lb). To a susvacuo. The residue was crystallized from acetonitrile to give 6.5 pension of Sa (10 g, 34.3 mmol) in 200 mL of water was added g (98.1%) of 7 b mp 195-197 "C; IR (KBr) 1720,1680,1610em-'. 5.0 g of concentrated HzS04. To the reaction mixture kept at 0 "C was gradually added a solution of NaN02 (2.6g, 37.7 mmol) Anal. (C19H24N404.Hz0) C, H, N. Method A. Ethyl l-Ethyl-l,4-dihydro-7-methoxy-6-nitro- in 3 mL of water while the temperature was kept at 3-6 "C. The 4-oxo-1,8-naphthyridine-3-carboxylate (sa). To a stirred resulting mixture was added a t room temperature to a solution mixture of concentrated HNO, (150mL) and concentrated H2SO4 of cuprous chloride (6.8g, 38.0 mmol) and KCN (8.6g, 132 mmol) in 160 mL of water. The reaction mixture was maintained at 80 (200mL) was added portionwise 50 g (0.181mol) of 7a at room "C for 1 h, basified with aqueous ammonia, and extracted with temperature. After complete addition, the reaction mixture was CHC1,. The extract was washed with water, dried, and concenheated at 60-70 "C for 1.5 h, poured onto 1.5 L of ice-water, and extracted with CHC13. The extract was washed with 7% NaHCO,, trated to dryness in vacuo. The residue was chromatographed dried, and concentrated in vacuo to give 47.0 g (80.8%) of 8a: IR on silica1 gel with CHCl, to give 6.8 g (65.8%) of llb: IR (KBr) 2200,1720,1630em-'; EIMS, m / z 301 (M'), 229 (M' - C2H4COz); (KBr) 1720,1610,1540,1360em-'; EIMS, m / z 321 (M'), 249 (M' lH NMR (CDCl,) 6 8.94 (1H, s, H-5), 8.58 (1 H, s, H-2), 4.42 (4 - CzH4C02). Method B. Ethyl l-Ethyl-1,4-dihydro-6-nitro-4-oxo-7-( 1H, q, J = 7 Hz, C02CH2CH3, NCH2CH,), 1.55 (3 H, t, J = 7 Hz, pyrrolidinyl)-1,8-naphthyridine-3-carboxylate(8c). To a CO,CHZCH,), 1.45 (3 H, t, J = 7 Hz, NCHzCH3). Method F. 6-Chloro-l-ethyl-1,4-dihydro-7-methoxy-4stirred solution of 8a (1.0g, 3.11 mmol) in 30 mL of acetonitrile was added 3 mL of pyrrolidine. The mixture was heated to reflux oxo-1,8-naphthyridine-3-carboxylicAcid (12a). A mixture for 3 h. The solvent was evaporated in vacuo. The residue was containing 3.0g (9.65mmol) of lla,80 mL of 1 N HCl, and 20 mL of EtOH was heated at 90-95 "C for 1 h. The resulting clear crystallized from AcOEt to give 1.03 g (91.6%) of 8c: IR (KBr) solution was allowed to cool. The precipitate was filtered off, 1720, 1680,1540, 1350 cm-'. washed with EtOH, and recrystallized to give 2.3 g (72.3%) of Met hod C. Ethyl 6-Amino-1-ethyl-l,l-dihydro-'l-met h12a: IR (KBr) 3000,1700,1600cm-'; EIMS, m / z 282 (M'), 238 oxy-4-oxo-l,8-naphthyridine-3-carboxylate (9a). To a stirred (M' - COP). solution of 8a (20 g, 62.3 mmol) in 400 mL of AcOH was added Method G. 6-Chloro-l-ethyl-7-hydroxy-1,4-dihydro-4portionwise 40 g of reduced-iron powder at such a rate that the temperature did not rise over 80 "C. After complete addition, oxo-1,8-naphthyridine-3-carboxylic Acid (13). A stirred suspension of lla (5.0g, 16.1 mmol) in 50 mL of 1 N NaOH was the reaction mixture was heated at 80 "C for 1 h and cooled to heated at 90-95 OC for 45 min. Neutralization of the mixture with room temperature. After addition of EtOH (500mL),the mixture was filtered to remove insoluble materials, and the filtrate was 30% AcOH afforded 4.1 g (94.7%) of 13: IR (KBr) 1705,1650, 1620 em-'. concentrated to dryness in vacuo. The residue was taken up in 1-ethyl-1,4-dihydro-4-oxo1,8800 mL of water and allowed to stand under ice cooling to give Met hod H. 6,7-Dichloronaphthyridine-3-carboxylicAcid (14). A mixture containing 14.5 g (79.7%) of Sa: IR (KBr) 3420,3280,2950,1680,1620 cm-'; 2.7 g (10mmol) of 13 and 15 mL of POCl, was heated to reflux 'H NMR (Me2SO-de)6 8.63 (1 H, s, H-2), 7.62 (1 H, s, H-5), 5.47 for 30 min. The excess of POCl, was evaporated in vacuo, and (2H,s,NHP),4.45and 4.23 (each 2 H , q , J = 7 Hz,NCH&H,, the residue was poured onto ice-water to give precipitates, which OCH2CH,), 4.07 (3 H, s, OCH,), 1.42 and 1.30 (each 3 H, t, J = were filtered off, washed with water, and recrystallized to give 7 Hz, NCH,CH,,OCH2CH&. 1.9 g (65.7%) of 14: IR (KBr) 1720 em-'; EIMS, m / z 287 (M'), 3-(Ethoxycarbony1)- 1-ethy 1-1,4-dihydro-7-met hoxy -4-oxo243 (M' - C02). 1,8-naphthyl'idine-6-diazonium Tetrafluoroborate (1Oc). To 6-Chloro-1-ethyl1,4-dihydro-4-oxo-7-( 1-pyrrolidiny1)1,8a stirred solution of Sa (5.0g, 17.2 mmol) in 65 mL of 42% HBF4 naphthyridine-3-carboxylic Acid (15a). (A)Method I. A was added dropwise a solution of NaN02 (1.2g, 17.4 mmol) in mixture containing 0.50 g (1.74mmol) of 14,0.75g (10.5mmol) 2 mL of water while the temperature was maintained at 5-10 "C. of pyrrolidine, and 20 mL of acetonitrile was heated to reflux for After 30 min, the precipitate was collected by filtration, washed 45 min. The solvent was evaporated in vacuo, and the residue with a mixture of MeOH-Et20 (1:l v/v), and crystallized from was taken up in 10 mL of water, neutralized with 30% AcOH, water to give 5.7 g (85.0%) of 1Oc: mp 148-149 "C dec; IR (KBr) and extracted with CHC1,. The extract was concentrated to 2200, 1720, 1620 em-'. Anal. (C14H15BF4N404) C, H, F, N. Thermal Decomposition of 1Oc. A mixture containing 2.0 dryness to give 0.48 g (85.7%) of 15a: IR (KBr) 1700,1610em-'; EIMS, m / z 321 (M'), 277 (M' - COz), 249 (M' - CZHdCO1). g (5.13 mmol) of 1Oc and 2.0 g of anhydrous Na2S04was heated a t 73-85 "C for 3 h. The reaction mixture was taken up in 80 According to this procedure, compounds 15b,c were prepared mL of EtOH-CHC1, (l:l,v/v) and filtered to remove insoluble from 14 with anhyrous piperazine and N-methylpiperazine, respectively. materials. The filtrate was concentrated to dryness in vacuo. The residue was chromatographed on silica gel with CHCls. The solid (B)Method J. A mixture containing 0.50 g (1.77 mmol) of resulting from the main fraction was recrystallized from n-hex12a,1.0 g (14.1 mmol) of pyrrolidine, and 30 mL of DMF was allowed to stir a t 50-60 "C for 2 h. After the same workup as in method I, 15a (0.43 g, 75.6%) was obtained. Method K. Ethyl 6-Cyano-l-ethyl-1,4-dihydro-4-oxo-7-( 1(19) Matsumoto, J.; Hirose, T.; Mishio, S., manuscript in preparapiperazinyl)-1,8-naphthyridine-3-carboxylate (17b). To a tion.
1,4-Dihydro-4-oxo-l,8-naphthyridine-3-carboxylic Acids
Journal of Medicinal Chemistry, 1984, Vol. 27,No. 3 299
Method Q. l-Ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(lrefluxed solution of anhydrous piperazine (2.0 g, 23.2 mmol) in piperazinyl)-l,S-naphthyridine-3-carboxylic Acid (Enoxacin, 30 mL of acetonitrile was added 1.0 g (3.27 mmol) of 16 over a 24b). A suspension of 20b (5.0 g, 12.8 mmol) in 50 mL of 10% period of about 15 min with stirring. The reaction mixture was HC1 was heated to reflux for 4 h and allowed to cool. The resulting heated to reflux for an additional 1 h and allowed to cool. The precipitate was filtered off and washed with EtOH to give 4.15 resulting precipitate was filtered off and recrystallized to give 0.60 g (91%) of 24beHC1. This salt was dissolved in 120 mL of hot g (51.7%) of 17b: IR (KBr) 2200,1710,1620 cm-'; EIMS, m / z water and treated with charcoal, and, after the addition of 20% 355 (M+), 283 (M+ - CzH4COZ). HCl(5 mL), the solution was cooled to give the pure hydrochloride In a similar manner, compounds 17a,c were prepared from 16 C, H, C1, F, N. of 24b,mp >300 "C. Anal. (C15H17FN403.HC1) with pyrrolidine and N-methylpiperazine, respectively. Ethyl 7-(4-Acetyl-l-piperazinyl)-l-ethyl-6-fluoro-l,4-di- The hydrochloride was dissolved again in 20 mL of water with heating. Subsequent addition of 2.2 mL of concentrated aqueous hydro-4-oxo-1,8-naphthyridine-3-carboxylate (20b). (A) ammonia under cooling led to precipitation of crystals, which were Method L. To a stirred solution of 9b (1.0 g, 2.56 mmol) in 20 filtered off and washed with cold water to give 3.1 g (75.3%) of mL of 1N HC1 was added dropwise a solution of NaNOz (250 24b IR (KBr) 2400-2000,1615 cm-'; EIMS, m/z 320 (M'), 276 mg, 3.6 mmol) in 3 mL of water, while the temperature was (M+- COJ, 234 (M+ - CHFNCH~); 'H NMR (Me2SO-d,) 6 8.92 maintained at 0-3 "C. After the reaction mixture was stirred for (1H, s, H-2), 8.00 (1H, d, J = 14 Hz, H-5), 4.49 (2 H, q, J = 7 5 min, aqueous 65% HPFs was added to the mixture until precipitation was complete. The precipitate was fdtered off and dried Hz, NCHzCH3), 3.76 and 2.85 (each 4 H, m, piperazine H), 1.39 over P2O5 in vacuo to give 1.3 g (94.2%) of the diazonium salt (3 H, t, J = 7 Hz, NCHZCH,). 19b: mp 128-131 "C dec; IR (KBr) 2180 cm-'. A suspension of 24b.AcOH: mp 228-229 "C (recrystallized from EtOH). Anal. 19b (1.0 g, 1.84 mmol) in 40 mL of n-heptane was heated to reflux (C15H17FN403~C2H402) C, H, F, N. 24b.MeS03H: mp >300 "C (recrystallized from EtOH-HzO). Anal. (C15H17FN403CH403S) for 15 min and allowed to cool. The resulting precipitate was C, H, F, N, S. collected by filtration, taken up in 30 mL of water, and extracted Method R. l-Ethyl-6-fluoro-1,4-dihydro-7-(4-methyl-lwith CHC13. The extract was dried and concentrated to dryness piperazinyl)-4-oxo-l,S-napht hyridine-3-carboxylicAcid (244. in vacuo to afford a crude product, which was recrystallized to A mixture containing 6.0 g (16.8 mmol) of 24b, 12 mL of 37% give 0.26 g (36.2%) of 14b IR (KBr) 1720,1680,1620cm-'; EIMS, m/z 390 (M'), 318 (M+ - C2H4COz);'H NMR (CDC13)6 8.55 (1 formalin, and 18 mL of formic acid was heated at 120-125 "C for H, s, H-2), 8.23 (1H, d, J = 14 Hz, H-5), 4.45 (2 H, q, J = 7 Hz, 17 h with stirring. The reaction mixture was concentrated to dryness in vacuo. The residue was taken up in 30 mL of water. COzCHzCH3),4.37 (2 H, q, J = 7 Hz, NCH2CH3),3.80 (8 H, s, piperazine H), 2.18 (3 H, s, COCHJ, 1.50 (3 H, t, J = 7 Hz, The mixture was adjusted at first to pH 9 with 10% NaOH and COZCHzCH3), 1.47 (3 H, t, J = 7 Hz, NCHzCH3). subsequently to pH 7 with AcOH and then extracted with CHCIB. Similarly prepared was compound 20a from 9c via 19a using The extract was washed with water, dried, and concentrated to cyclohexane. Diazonium salt 19a: mp 125-130 "C dec; IR (KBr) dryness in vacuo. The residue was crystallized from CHZClz-EtOH 2180 cm-l. to give 5.0 g (75%) of 24c: IR (KBr) 1710,1630 cm-'; EIMS, m/z (B)Method M. A mixture containiig 3.62 g (10 mmol) of ethyl 334 (M'), 290 (M' - COZ), 220 [M+ - CH&H,N(CH,)=CH]; 'H 7-(4-acetyl-l-piperazinyl)-6-fluoro-1,4-dihydro-4-0~0-1,8-NMR (CDC13 6 8.78 (1H, s, H-2), 8.13 (1H, d, J = 14 Hz, H-5), 4.48 (2 H , q , J = 7 Hz,NCHzCH3),3.95 (4 H, m, CHzNCHz),2.60 naphthyridine-3-carboxylate (21b),152.76 g (20 mmol) of KzCO3, and 40 mL of DMF was heated at 100 "C for 30 min with stirring. (4 H, m, CH2N4CHz),2.39 (3 H, s, NCH3), 1.52 (3 H, t, J = 7 Hz, To this mixture was added 4.68 g (30 mmol) of ethyl iodide. The NCHzCH3), 14.2 (1 H, br s, COOH). l-Ethyl-6-fluoro-1,4-dihydro-7hydroxy-4-oxo-1,Sresulting mixture was allowed to stir a t the same temperature naphthyridine-3-carboxylic Acid (25). A suspension of 20b for 3 h and filtered to remove insoluble materials. The filtrate was concentrated to dryness in vacuo. The residue was taken up (7.8 g, 20 mmol) in 100 mL of 20% NaOH was heated to reflux in a mixture of water (30 mL) and CHC13 (50 mL). The organic for 43 h with stirring. After ice cooling, the precipitate (the phase was separated, washed with water, dried, and chromatodisodium salt of 25) was filtered off and taken up in 200 mL of graphed on silica gel with CHC13to give 3.5 g (89.7%) of 20b this hot water. The aqueous solution was treated with charcoal, compound was identical in all respects with an authentic specimen acidified with 20 mL of 10% HC1, while hot, and allowed to cool of 20b prepared by the method L. to give precipitates, which were fitered off and washed with water. Also prepared according to this procedure were 20a [from 21a15 Recrystallization from water-EtOH (1:4, v/v) gave 4.13 g (82.0%) with ethyl iodide (Table I)] and 31 and 33 [from 21b with ethylene of 25: mp >300 "C; IR (KBr) 1710 cm-'. Anal. (CllH9FN204) bromohydrin and 2-fluoroethyltosylate, respectively (Table III)]. C, H, F, N. Method N. 1-Ethy1-1,4-dihydro-6-nitro-4-oxo-7-(1- 7-Chloro-l-ethyl-6-fluoro-1,4-dihydro-4-oxo-l,Spiperazinyl)-l,S-naphthyridine-3~carboxylic Acid (22b). A naphthyridine-3-carboxylicAcid (26). A mixture containing mixture containing 4.1 g (9.82 mmol) of Sb,40 mL of 15% HC1, 9.5 g (37.7 mmol) of 25 and 50 mL of POC13 was heated to reflux and 20 mL of EtOH was heated at 90-100 "C for 100 min with for 15 min. The excess of POC13 was evaporated in vacuo, and stirring and allowed to cool. The resulting precipitate (the HC1 the residue was poured into 300 mL of ice-water. After 30 min salt of 22b) was filtered off and dissolved in a minimum volume of stirring, the precipitate was filtered off and washed successively of hot water. The aqueous solution was basified with aqueous with water and acetone to give 9.3 g (91.4%) of 26: mp 265-267 ammonia to afford precipitates, which were filtered off and re"C (recrystallized from CH3CN);IR (KBr) 1720 cm-l; EIMS, m/z crystallized to give 2.76 g (80.9%) of 22b: IR (KBr) 1630, 1600, 270 (M'), 226 (M+ - COz);'H NMR (CDC13) 6 8.98 (1H, s, H-2), 1560, 1360 cm-'; EIMS, m/z 347 (M+), 303 (M+ - COz). 8.56 (1H, d, J = 8 Hz, H-5), 4.62 (2 H, q, J = 7 Hz, NCHzCH3), Method 0. 6-Amino-l-ethyl-1,4-dihydro-4-oxo-7-(l- 1.58 (3 H, t, J = 7 Hz, NCHzCH3), 14.2 (1H, s, COOH). Anal. pyrrolidinyl)-l,8-naphthyridine-3-carboxylic Acid (23a). A (CllH&lFNZ03) C, H, C1, F, N. stirred suspension of 9c (0.90 g, 2.72 mmol) in 20 mL of 1N NaOH Method S. 7-Amino-l-ethyl-6-fluoro-1,4-dihydro-4-oxowas heated at 80-90 "C for 15 min. The resulting solution was l,S-naphthyridine-3-carboxylic Acid (27a). To a stirred sotreated with charcoal and neutralized with AcOH to give 0.70 g lution of 26 (1.35 g, 5 mmol) in 20 mL of DMF was added 4.8 g (85.1%) of 23a: IR (KBr) 3360,3300,1680,1610 cm-'; EIMS, (50 mmol) of ammonium carbonate. The mixture was heated at m / z 302 (M+), 258 (M+ - COz). 90 "C for 2 h, after which time an additional 2.4 g (25 mmol) of Method P. 6-Amino-l-ethyl-1,4-dihydro-7-(4-methyl-l-ammonium carbonate was added. The reaction was allowed to piperazinyl)-4-oxo-1,&naphthyridine-3-carbxylic Acid (23c). run for an additional 1.5 h. After the mixture was cooled, the A mixture containing 0.50g (1.36 -01) of 22c,0.10 g of 5% Pd/C, precipitate was filtered off, washed with water, and recrystallized and 50 mL of EtOH was hydrogenated at 50-60 "C until the to give 1.1 g (87.6%) of 27a (Table 11): EIMS, m/z 251 (M+), required volume of Hz had been taken up. The reaction mixture 207 (M+ - COZ). was fitered to remove the catalyst. The filtrate was concentrated Method T. l-Ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(3-oxo-lto dryness in vacuo. The crystalline residue was purified by piperazinyl)-l,S-naphthyridine-3-carboxylic Acid (27c). A column chromatography on silica gel with CHCl, to give 0.12 g mixture containing 1.5 g (5.55 mmol) of 26, 1.11g (11.1 mmol) (26.1%) of 23c: EIMS, m/z 331 (M+), 287 (M+ - COz), 261 [M+ of 2-oxopiperazine, and 100 mL of acetonitrile was heated to reflux - CH~=N(CH~)CH&HS], 217 [287 - C H Z N ( C H ~ ) C H ~ C H ~ ] . for 1 h and allowed to cool. The precipitate was filtered off,
300 Journal of Medicinal Chemistry, 1984, Vol. 27, No. 3 washed with water, and recrystallized from DMF-EtOH (ca. 1:1, v/v) to give 1.35 g (71.4%) of 27c: IR (KBr) 3400,3200,3050, 2400,1710,1670,1620 cm-l; EIMS,m/z 334 (M+),290 (M+- Cod.
Matsumoto et al.
off, washed with water, and taken up in ca. 40mL of 10% AcOH with heating. The solution was adjusted to pH 9 with aqueous ammonia and cooled to give precipitates, which were dissolved Method U. l-Ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(4-n in 10% AcOH; the solution was neutralized with ammonia to give propyl-l-piperazinyl)-l,8-naphthyridine-3-carboxylic Acid 2.90g (82.4%) of 36: IR (KBr) 1620cm-'; 'H NMR (Me2SO-d6) (28a). A mixture containing 1.0g (3.12mmol) of 24b,5 mL of 6 8.82(1 H, 8, H-2), 8.05(1 H, d, J = 14Hz, H-5),7.76(1 H, dd, propionaldehyde, and 5 mL of formic acid was heated at 90 OC J = 16 and 9 Hz, NCH=CH2), 5.82(1 H, dd, J = 16 and 2 Hz, for 17 h with stirring. The residue was taken up in 20 mL of 10% NCH=CH trans), 5.40(1 H, dd, J = 9 and 2 Hz, NCH=CH cis), HCl and extracted with three 20-mL portions of CHC13. The 3.75and 2.84 (8H, piperazine H). combined extracts were dried and concentrated to dryness. The The initial filtrate and the washings were combined and conprecipitate was recrystallized from EtOH-CH2C12to give 0.02g centrated to dryness. Recrystallization from an ca. 1:l EtOHof 29 which was identical with its authentic specimen prepared CHC13 mixture gave 0.15g (5.4%) of 6-fluoro-1,4-dihydro-7hydroxy-4-oxo-l-vinyl-l,8-naphthyridine-3-carboxylic acid by method V. The foregoing acidic aqueous layer was adjusted to pH 7with 10% NaOH and extracted with three 20-mL portions (41): mp 256-259 "C; IR (KBr) 1700cm-'; 'H NMR (Me2SO-d6) 6 8.93(1 H, 8, H-2), 8.25(1H, d, J = 10Hz, H-5), 7.73(1 H, dd, of CHC13. The combined extracts were dried, and the solvent J = 16 and 9 Hz, NCH-CHJ, 5.93(1 H, dd, J = 16 and 2 Hz, was evaporated. The residue was crystallized from CHzClz-EtOH (ca. 1:1,v/v) to give 0.51 g (46.5%) of 28a. NCH=CH trans), 5.53(1 H, dd, J = 9 and 2 Hz, NCH==CH cis), Also prepared according to this procedure were 28b,c(Table -13.0 (br, COOH and/or OH). Anal. (CllH7FN204)C, H, F, 11) from 24b using n-butyraldehyde and sec-butyraldehyde, reN. In Vitro Antibacterial Activity. According to the method spectively. Method V. l-Ethyl-6-fluoro-7-(4-formyl-l-piperazinyl)- of Goto et a1.,20the MIC (in micrograms per milliliter) was determined by the twofold agar dilution method using Mueller1,4-dihydro-4-oxo-l,8-naphthyridine-3-carboxylic Acid (29). Hinton agar (pH 7.4,Difco); bacterial inocula contained apA mixture containing 5.0 g (15.6mmol) of 24b,50 mL of formic acid, and 10mL of formamide was heated to reflux for 2 h. After proximately lo6 colony-forming units and the bacterial growth was observed after 20-h incubation at 37 "C. the mixture was cooled, the resulting precipitate was filtered off In Vivo Efficacy on SystemicInfections. In vivo activity and washed with acetone to give 5.3g (97.5%) of 29: IR (KBr) 1720,1670,1625cm-l. assay was carried out according to the method of Shimizu et ala2' Method W.Ethyl 7-(4-Acetyl-l-piperazinyl)-l-(2-chloro- Groups of 10or more male mice (Std-ddY, 20 f 2g) were infected ethyl)-6-fluoro-1,4-dihydro-4-oxo1,8-naphthyridine-3- with Staphylococcus aureus 50774 (iv, 5 X lo8cells), Escherichia coli P-5101(ip, 9 X lo6 cells), and Pseudomonas aeruginosa 12 carboxylate (32). To a stirred solution of 31 (5.4g, 13.3mmol) (ip, 4 X lo3cells). The test compounds were suspended in 0.2% in 60 mL of CHC13 was added a solution of SOClz (3.17g, 26.6 sodium carboxymethylcellulose and administered orally at 0- and mmol) in 10mL of CHC13. The mixture was heated to reflux for 6-h postinfection. Survival rates were evaluated after 2 weeks 30 min, cooled, mixed with 30 mL of water, and neutralized with for the staphylococcal infection and after 1 week for others. saturated NaHC03. The organic layer was separated, washed with Acute Toxicity Test. A suspension of the test compound in water, dried, and concentrated to dryness. The residual solid was 0.2% carboxymethylcellulosein different concentrations was orally chromatographed on silica gel with CHC13 to give 5.1g (90.3%) given to male mice (Std-ddY, four to eight in each group) at a of 32: EIMS, m / z 424 (M'), 379 (M+ - C2H50), 352 (M+ dose of 0.1 mL/10 g of body weight. The number of dead mice CzH4COz);IR (KBr) 1720,1620cm-*. Method X. Ethyl 7-(4-Acetyl-l-piperazinyl)-6-fluoro-l- was counted after 7 days, and the LDso value (milligrams per (difluoromethyl)-l,4-dihydro-4-oxo-l,8-naphthyridine-3- kilogram) was calculated in according to the Behrens-Kaerber method.22 carboxylate (34). To a refluxed solution of 21b (10.8g, 30 mmol) in 300 mL of DMF was added dropwise a solution of sodium Acknowledgment. The authors are grateful to Dr. M. chlorodifluoroacetate (11.4g, 75 mmol) in 30 mL of DMF over Shimizu for his encouragement throughout this work. a period of 5 min. The mixture was allowed to reflux for an Thanks are also due to Drs.Y. Takase and S. Nakamura additional 30 min and then cooled. The insoluble material was fiitered off and washed with CHzClzto give 8.5g (78.7% recovery) for the biological test, H. Okada for his assistance in the of the unchanged compound 21b. The filtrate and the washings synthetic work, and members of the analytical section of were combined and concentrated to dryness in vacuo. The residue these laboratories for elemental analyses and spectral was triturated with hot CHCl,. The CHC13 solution was washed measurements. with water and then dried over KzCO3, and the solvent was evaporated to leave a crude product, which was chromatographed Registry No. 3a,57948-69-3; 3b,54132-31-9; 3c,54132-45-5; on silica gel with CHC1,; the main fraction gave 2.4 g (19.4%) of 7a,76105-70-9; 7b,54132-41-1; 8a,79286-67-2; 8b,87939-04-6; 8c, 34: IR (KBr) 1730,1650,1625cm-l; EIMS, m / z 412 (M'), 340 65095-80-9; 9a,79286-68-3; 9b,87938-96-3; 9c, 70186-25-3; lOc, (M+ - C,H,CO,); 'H NMR (MezSO-d6)6 8.57(1H, s, H-2), 7.96 79286-70-7; lla,87938-89-4;llb,87938-90-7; 12a,87938-91-8;12b, (1H, d, J = 14Hz, H-5), 8.35(1 H, t, J 59 Hz, CHFZ), 4.26(2 87939-18-2; 13,87938-92-9; 14,70186-28-6; 15a,70186-29-7; 15b, H, q, J = 7 Hz, NCH2CHB),3.81 and 3.36 (8H, piperazine H), 87938-93-0; 15c,87938-94-1; 16,75064-88-9; 17a,87938-95-2; 17b, 2.06 (3 H, 8 , COCHS), 1.29 (3 H, t, J = 7 Hz, NCH&H3). 76105-58-3; 17c,76105-72-1; 18a,76105-57-2; 18b,87939-19-3; 18c, l-piperazinyl)-6-fluoro1,4-diMethod Y. 7 44-Acetyl76105-73-2; 19a,87939-01-3; 19b,87938-98-5; 20a,87938-99-6; 20b, hydro-4-oxo-l-vinyl-l,8-naphthyridine3-carboxylic Acid (35). 82671-13-4; 21a,87939-02-4; 21b,84233-60-3; 22a,87939-20-6; 22b, To a hot solution of 32 (2.13g, 5 mmol) in 15 mL of EtOH was 74274-57-0; 22b-HC1,87939-05-7; 2213,87939-08-0; 23a,87939-06-8; added a solution of KOH (0.84g, 15 mmol) in 15 mL of EtOH. 23b,74274-56-9; 23c,87939-07-9; 24a,74274-63-8; 24b,74011-58-8; The reaction mixture was heated to reflux for 2 h and allowed 24b.HC1, 74011-31-7; 24b*AcOH, 75167-07-6; 24b.MeS03H, to cool. The precipitate was filtered off, washed with EtOH, and 75167-06-5; 24c,74274-66-1; 25,87939-09-1; 25.2Na,87939-10-4; dissolved in 20 mL of boiling water. The aqueous solution was 26,79286-73-0; 27a,87939-11-5; 27b,87939-21-7; 27c,87939-12-6; treated with charcoal and adjusted to pH 4-5 with 10% AcOH 27d,74274-65-0; 27e,74274-64-9; 27f,87939-22-8; 27g,74274-60-5; to give 1.45 g (80.6%) of 35: IR (KBr) 1710 cm-'; 'H NMR 27h,87939-23-9; 27i,87939-24-0; 27j,87939-25-1; 27k,7427470-7; (Me@O-ds) 6 8.90(1 H, s, H-2), 8.14(1H,d, J = 13.5Hz, H-5), 271,74274-67-2; 28a,74274-68-3; 28b,74274-69-4; 28c,87939-14-8; 7.83(1 H, dd, J = 16 and 9 Hz, NCH=CH2), 5.88(1 H, dd, J = 2 and 16 Hz, NCH=CH trans), 5.45 (1 H, dd, J = 2 and 9 Hz, NCH=CH cis), 3.78(8H, piperazine H), 2.03 (3H, s, COCHB), (20) Goto, S.;Jo, K.; Kawakita, T.; Kosaki, N.; Mitsuhashi, S.; 14.95 (1 H, br s, COOH). Nishino, T.; Ohsawa, N.; Tanami, H. Chemotherapy 1981,29, Method Z. 6-Fluoro-1,4-dihydro-4-oxo-7-(176. piperazinyl)-l-vinyl-1,8-naphthyridine-3-carboxylic Acid (21) Shimizu, M.; Takase, Y.; Nakamura, S.; Katae, H.; Minami, A.; (36). A stirred suspension of 35 (4.0g, 11.1mmol) in 20 mL of Nakata, K.; Kurobe, N. Antimicrob. Agents Chemother. 1976, 10% NaOH was heated to reflux for 2 h, allowed to cool, and 9,569. (22) Kaerber, G.Arch. Exp. Pathol. Pharmacol. 1931,162,480. adjusted to pH 7with 30% AcOH. The resulting solid was fiitered
J.Med. Chem. 1984,27, 301-306 29,87939-13-7; 30,78903-83-0; 31,87939-03-5; 32,87939-15-9; 33, 84209-43-8; 34,87939-16-0; 35,87939-17-1; 36,74274-71-8; 36.HC1, 75167-167; 36-MeSO3H,75167-17-8; 37,7427472-9; 38,84209-347; 39, 84209-33-6; 40, 87939-26-2; 41, 84424-27-1; ethyl 1,4-di87938hydro-7-methoxy-4-oxo-1,8-naphthyridine-3-carboxylate, 88-3;ethyl 7-chloro-l-ethyl-4-oxo-1,&naphthyridine-3-carboxylate, 56654-05-8; sec-butyraldehyde, 78-84-2; N-methylpiperazine, 109-01-3; N-acetylpiperazine, 13889-98-0; pyrrolidine, 123-75-1;
301
piperazine, 110-85-0; ethylene bromohydrin, 540-51-2; 2-fluorethyl tosylate, 383-50-6; 2-oxopiperazine, 5625-67-2;propionaldehyde, 123-38-6; butyraldehyde, 123-72-8; sodium chlorodifluoroacetate, 1895-39-2; ethylenediamine, 107-15-3; piperidine, 110-89-4; morpholine, 110-91-8; thiomorpholine, 123-90-0; homopiperazine, 505-66-8; azepine, 12764-48-6; azocine, 1121-92-2; l-phenylpiperazine, 92-54-6; 1-benzylpiperazine, 2759-28-6; l-ethylpiperazine, 5308-25-8.
Structure-Activity Relationships of Sparsomycin and Its Analogues. Octylsparsomycin: The First Analogue More Active than Sparsomycin Rob M. J. Liskamp,? J. Hans Colstee,? Harry C. J. Ottenheijm,*J Peter Lelieveld,’ and Wil Akkerman’ Department of Organic Chemistry, University of Nijmegen, Toernooiveld, 6525 E D Nijmegen, and Radiobiological Institute TNO, Lange Kleiweg 151 2288 GJ Rijswijk, The Netherlands. Received November 12,1982 Nine analogues of sparsomycin (1) were synthesized, and their cytostatic activity was studied in an in vitro clonogenic L1210 assay by measuring the inhibition of colony formation. The activity of an analogue, expressed as an ID,, value, was compared to that of sparsomycin (Table I). Each analogue possesses not more than two structural modifications of the sparsomycin molecule 1. Comparison of the activity of 1 with that of the stereomers 2-4, having RcSS, ScSs, and RcRs chirality, respectively, shows that the S configuration of the chiral carbon atom is essential for an optimal activity, whereas the R chirality of the sulfoxide sulfur atom of sparsomycin is of importance. Study of the IDm values of the S-deoxo analogues 10 and 11, as well as the compounds 14 and 15 having the 0-sulfoxide function, indicate that the presence of an oxygen atom on the a-sulfur atom is essential. Isomerization of the trans double bond into the cis double bond yields isosparsomycin (16, Scheme 11), which has a low activity. The cytostatic activity of sparsomycin seems to be related to its lipophilicity: octylsparsomycin (19) was shown to be three times as effective as sparsomycin.
The development of a flexible synthesis is a prerequisite for thorough studies on the biological activity and/or biochemical mechanism of the interaction of a molecule. An which this view, is sparsomycin
(0.l
(1) Sparsomycin is a metabolite of Streptomyces (Argoudelis,A. D.; Herr, R. R. Antimicrob. Agents Chemother.
1962, 780) and Streptomyces cuspidosporus (Higashide, E.; Hasegawa, T.; Mizuno, K.; Akaide, H. Takeda Kenkyusho Nempo. 1966,25, 1; Chem. Abstr. 1967,66, 54328). (2) Ottenheijm, H. C. J.; Liskamp, R. M. J.; Tijhuis, M. W. TetM. J.; vanLett. rahedron Nispen, 1979, S. P. 387. J. M.; Ottenheijm, Boots, H.H. A.;C.Tijhuis, J.; Liskamp, M. W. R. J.
HN~NLs~w%e A H (vg
H
0
transformed5J1 and/or virus infected cells12),and various cell-free systems.13 The behavior of sparsomycin with
I;*
Org. Chem. 1981,46, 3273. (3) Liskamp, R. M. J.; Zeegers, H. J. M.; Ottenheijm, H. C. J. J . Org. Chem. 1981,46, 5408. 1 : Sparsornycin I SC -Rsl (4) Heluuist, P.: Shekhani, M. S. J. Am. Chem. SOC. 1979, 101. 1057. This antibiotic has been synthesized only r e ~ e n t l y . ~ - ~ (5) Lin, C.-C. L.; Dubois, R. J. J. Med. Chem. 1977,20, 337. Consequently, the structure-activity relationship studies (6) Vince, R.; Brownell, Lee, C. K. Biochem. Biophys. Res. Commun. 1977,75,563. Lee, C. K.; Vince, R. J. Med. Chem. 1978, of 1 that have appeared so fars7 concern analogues in 21, 176. which several structural parameters have been varied si( 7 ) Dubois, R. J.; Lin, C X . L.; Michel, B. L. J . Pharm. Sci. 1975, multaneously, thus allowing only a limited interpretation 64, 825. of the results with regard to the role of the various (8) Pestka, S. Annu. Rev. Microbiol. 1971,25, 488. Vazquez, D. structural fragments. FEBS Lett. 1974, 40, ,963. Vazquez, D. Mol. Biol. Biochem. The interpretaton and comparison of the available inBiophys. 1979, 30. (9) Slechta, L. In “Antibiotics I”; Gottlieb, D.; Shaw, P. D., Eds.; formation on structure-activity relationships of sparsoSpringer Verlag: New York, 1967; p 410. Bannister, R. E.; mycin encounters a second difficulty; the biological activity Hunt, D. E.; Pitillo, R. F. Can. J. Microbiol. 1967, 595. of the analogues has been determined in different systems (10) Goldberg, I. H.; Mitsugi, K. Biochem. Biophys. Res. Commun. (in vitro: KB cell culture5 and cell-free ribosomal systems6; 1966,23,453. Contreras, A.; Vazquez, D.; Carrasco, L. J. Anin vivo: P-388 system6 and Walker 256 system7). tibiot. 1938, 31,598. Gupta, R. S.; Siminovitch, L. BiochemSparsomycin (1) is a strong inhibitor of protein bioistry 1977, 16, 3209. synthesis and has therefore attracted widespread attention. (11) (a) Owen, S. P.; Dietz, A.; Camiener, G. W. Antimicrob. Agents Chemother. 1962, 772. (b) Kuwano, M.; Takenaka, K.; Ono, There is ample evidences that sparsomycin has its site of M. Biochim. Biophys. Acta 1979,563,479. (c) Bhuyan, B. K.; interaction in the large ribosomal subunit, where it preScheidt, L. G.; Fraser, I. J. Cancer Res. 1972, 32, 398. vents peptide transfer by interfering with the peptidyl(12) Contreras, A.; Carrasco, L. J. Virol. 1979,29, 114. Thiry, L. transferase center. Sparsomycin manifests its action in J. Gen. Virol. 1968, 2, 143. intact prokaryotic cells,9 eukaryotic cellslO (including (13) Baglioni, C. Biochim. Biophys. Acta 1966, 129, 642. Emmerich, B.; Hoffmann, H.; Erben, V.; Rastetter, J. Eiochim. Biophys. Acta 1976, 44, 460. Pestka, s. Proc. Natl. Acad. Sci. University of Nijmegen. U.S.A. 1968, 61, 726. Carrasco, L. Fresno, M.; Vazquez, D. Radiobiological Institute. FEES Lett. 1975, 52, 236. H
-
*
0022-2623/84/1827-0301$01.50/0 0 1984 American Chemical Society
